Lesort M, Attanavanich K, Zhang J, Johnson G V
Department of Psychiatry and Behavioral Neurobiology, University of Alabama at Birmingham, Birmingham, Alabama 35294-0017, USA.
J Biol Chem. 1998 May 15;273(20):11991-4. doi: 10.1074/jbc.273.20.11991.
Tissue transglutaminase is a calcium-dependent transamidating enzyme that has been postulated to play a role in the pathology of expanded CAG repeat disorders with polyglutamine expansions expressed within the affected proteins. Because intranuclear inclusions have recently been shown to be a common feature of many of these codon reiteration diseases, the nuclear localization and activity of tissue transglutaminase was examined. Subcellular fractionation of human neuroblastoma SH-SY5Y cells demonstrated that 93% of tissue transglutaminase is localized to the cytosol. Of the 7% found in the nucleus, 6% copurified with the chromatin-associated proteins, and the remaining 1% was in the nuclear matrix fraction. In situ transglutaminase activity was measured in the cytosolic and nuclear compartments of control cells, as well as cells treated with the calcium-mobilizing agent maitotoxin to increase endogenous tissue transglutaminase activity. These studies revealed that tissue transglutaminase was activated in the nucleus, a finding that was further supported by cytochemical analysis. Immunofluorescence studies revealed that nuclear proteins modified by transglutaminase exhibited a discrete punctate, as well as a diffuse staining pattern. Furthermore, different proteins were modified by transglutaminase in the nucleus compared with the cytosol. The results of these experiments clearly demonstrate localization of tissue transglutaminase in the nucleus that can be activated. These findings may have important implications in the formation of the insoluble nuclear inclusions, which are characteristic of codon reiteration diseases such as Huntington's disease and the spinocerebellar ataxias.
组织转谷氨酰胺酶是一种钙依赖性转酰胺酶，据推测它在受影响蛋白质中表达多聚谷氨酰胺扩展的CAG重复扩张性疾病的病理过程中发挥作用。由于最近已表明核内包涵体是许多这类密码子重复疾病的共同特征，因此对组织转谷氨酰胺酶的核定位和活性进行了研究。对人神经母细胞瘤SH-SY5Y细胞进行亚细胞分级分离表明，93%的组织转谷氨酰胺酶定位于细胞质中。在细胞核中发现的7%中，6%与染色质相关蛋白共纯化，其余1%存在于核基质部分。在对照细胞以及用钙动员剂 maitotoxin处理以增加内源性组织转谷氨酰胺酶活性的细胞的细胞质和细胞核区室中测量原位转谷氨酰胺酶活性。这些研究表明组织转谷氨酰胺酶在细胞核中被激活，这一发现得到了细胞化学分析的进一步支持。免疫荧光研究表明，经转谷氨酰胺酶修饰的核蛋白呈现离散的点状以及弥漫性染色模式。此外，与细胞质相比，细胞核中转谷氨酰胺酶修饰的蛋白质不同。这些实验结果清楚地证明了组织转谷氨酰胺酶在可被激活的细胞核中的定位。这些发现可能对不溶性核内包涵体的形成具有重要意义，而不溶性核内包涵体是亨廷顿舞蹈病和脊髓小脑共济失调等密码子重复疾病的特征。
